In recent years, regenerative medicine employing cell sheet engineering techniques has been attracting attention, and a large number of attempts to graft various types of cell sheets have been carried out. When grafting, a cell sheet is used in methods in which a single layer sheet is grafted, a uniform tissue formed by layering identical cell sheets is grafted, a tissue having a layered structure formed by layering several different types of cell sheets is grafted, etc. 20 to 30 types of cell sheets are currently produced, and progress has been made in clinical applications employing epithelial cell systems such as those of the cornea, retina, skin, the bladder epithelium, and the periodontal ligament.
However, there is a problem with the production of a cell sheet. The problem is that when peeling off cultured cells or a cell sheet from a culture dish, since cells adhere strongly to an adhesion protein on the culture dish, it is necessary to use a proteolytic enzyme, and as a result cells and extracellular matrix are damaged. Recently, a method that does not use a proteolytic enzyme, that is, a method for recovering a cell sheet in which a culture dish surface is coated with an N-isopropylacrylamide polymer (PIPAAm), which is a polyacrylamide for which the surface properties change depending on the temperature, has been developed. PIPAAm is a material whose affinity for water changes greatly on either side of 32° C. When this PIPAAm is fixed to a culture dish surface at a uniform nano-order thickness, the surface becomes hydrophobic at 37° C., which is suitable for culturing cells, and the cells adhere and proliferate. On the other hand, when the temperature is decreased to 32° C. or below after culturing, the surface becomes hydrophilic, and sheet-form cells can be peeled off without being damaged (ref. e.g. Patent Documents 1 and 2).
However, although an acrylamide polymer like PIPAAm does not itself have toxicity such as that shown by an acrylamide monomer, there is a possibility that, in the production process for an acrylamide polymer, unpolymerized acrylamide monomer, which has toxicity, will remain within the polymer, and this problem has been pointed out.